PROJECT SUMMARY Obesity and its complications including non-alcoholic fatty liver disease (NAFLD) have reached epidemic proportions worldwide. NAFLD is estimated to affect 30% of the population and is one of the leading causes of abnormal liver function in Western countries. NAFLD covers a wide spectrum of liver pathology ranging from a simple accumulation of fat to a more serious condition known as non-alcoholic steatohepatitis (NASH). Inflammation in the liver is a key process in the initiation, maintenance, and progression of NAFLD. However, the mechanisms triggering this inflammatory process remains unclear. B lymphocytes are central mediators of autoimmune and inflammatory disease because of their ability to secrete harmful substances. We have evidence that pro-inflammatory B cells accumulate in the liver of mice in a model of diet-induced NAFLD that is relevant to the human condition. The liver is a unique organ where immune cells interact with blood from the gastrointestinal tract that contains bacterial products that originate from the gut microbiota. During NAFLD, changes in the amounts and composition of the gut microbiota can lead to the leakage of bacterial products that promote inflammation. However, the role of B cells in the progression of NAFLD and the factors influencing their activation remain to be investigated. Our long-term goal is to reveal innovative mechanisms by which cells of the immune system promote NAFLD. As our preliminary data show that the liver accumulates pathogenic B cells in a mouse model of NAFLD, we plan to investigate their role in the pathogenesis of NAFLD. The central hypothesis is that intrahepatic B cells fuel local inflammation and fibrosis, resulting in the progression of NAFLD. We expect that hepatic B cell pathogenicity during NAFLD is supported by critical factors such as the entry of bacterial products from the intestines. Our specific aims are to identify the mechanisms by which hepatic B cells promote NAFLD (Aim 1), determine the intestinal-derived microbial factors fueling hepatic B cell pathogenicity (Aim 2), and reveal the mechanisms of B cell-mediated activation of hepatic stellate cells (Aim 3). It is well established that B lymphocytes play important roles in classical autoimmune disorders, and it is becoming increasingly clear that they contribute to tissue inflammation during metabolic disease. As NAFLD has no approved therapies for its treatment, the study of B cell effector and regulatory functions will provide mechanistic insights that can lead to new disease markers and therapeutics.